Nouri Mehraneh, Sharif Samaneh, Soltani Ehsan, Mozaffari-Jovin Sina, Abbaszadegan Mohammad Reza
Medical Genetics Research Center, Mashhad University of Medical Sciences, Azadi Square, Mashhad, Iran.
Department of Medical Genetics, Faculty of Medicine, Mashhad University of Medical Sciences, Mashhad, Iran.
J Transl Med. 2025 Jul 2;23(1):730. doi: 10.1186/s12967-025-06710-0.
Liquid biopsy is a minimally invasive method, which is an alternative to tissue biopsy in solid cancers to detect driver mutations, monitor treatment responses, and estimate prognosis. Exosomes are a group of extracellular vesicles that are detected in body fluids and are more stable than cell-free DNA and circulating tumor cells. Genetic data of parental cells can be packaged into exosomes, analysis of which could help determine the genetic modifications of cells before the emergence of disease symptoms. Here, we sought to investigate the KRAS mutation detection in exosomal DNA extracted from the plasma of patients with colorectal cancer.
This study was conducted on 42 colorectal cancer patients with stage I-III. First, exosomes were purified from the plasma of patients using Size Exclusion Chromatography (SEC), and then DNA was extracted from the samples using the phenol-chloroform-isoamyl alcohol (PCI) method. Next, the presence of three KRAS variants was examined using the Intplex quantitative PCR method.
The mutant allele frequency was measured with a sensitivity of 0.01%. About 85% of the tested exosomal DNA samples had one or two KRAS mutations. The measured mutant allele frequency had a median of 1.18% (with a range of 0.01-63.33%). Moreover, we evaluated the concordance of mutation detection between patient exosome and tumor tissue DNA samples.
We concluded that the investigation of mutation in genetic contents of exosomes extracted from the plasma of cancer patients at early stages could be performed with high sensitivity and could accelerate determination of the best therapy process for patients.
液体活检是一种微创方法，是实体癌组织活检的替代方法，可用于检测驱动突变、监测治疗反应和评估预后。外泌体是在体液中检测到的一组细胞外囊泡，比游离DNA和循环肿瘤细胞更稳定。亲代细胞的遗传数据可被包装到外泌体中，对其进行分析有助于在疾病症状出现之前确定细胞的基因修饰。在此，我们试图研究从结直肠癌患者血浆中提取的外泌体DNA中的KRAS突变检测。
本研究对42例I-III期结直肠癌患者进行。首先，使用尺寸排阻色谱法（SEC）从患者血浆中纯化外泌体，然后使用苯酚-氯仿-异戊醇（PCI）法从样品中提取DNA。接下来，使用Intplex定量PCR法检测三种KRAS变体的存在。
测量的突变等位基因频率灵敏度为0.01%。约85%的测试外泌体DNA样本有一个或两个KRAS突变。测量的突变等位基因频率中位数为1.18%（范围为0.01-63.33%）。此外，我们评估了患者外泌体和肿瘤组织DNA样本之间突变检测的一致性。
我们得出结论，对癌症患者早期血浆中提取的外泌体遗传内容物中的突变进行检测具有高灵敏度，并且可以加速为患者确定最佳治疗方案的过程。
